Important electrophysiological, pharmacological, and biochemical changes occur in myocardial cells during development of the heart, which obviously affect its functional properties. The identity and properties of the currents in young embryonic animals is not well understood. Young embryonic chick hearts have slowly-rising, Na-dependent, TTX-insensitive action potentials. The number of fast Na channels increases during development so that the old embryonic chick heart has a typical fast-rising Na-dependent action potential. Thus, the types and number of channels changes during development. Channel properties and kinetics may also change during development (e.g., K channels display less inward-rectification in young vs. old embryonic chick hearts). The identity and properties of the various cardiac channels will be examined at different stages of development, including changes in the Na channels using photoaffinity probes of tetrodotoxin. In these studies, whole-cell voltage clamp and patch clamp techniques will be used in single cells of embryonic chick hearts at different stages of development. In the adult heart, cAMP-dependent phosphorylation regulates the function of slow Ca channels (and perhaps other channels). Phosphorylation by other protein kinases may also regulate channel function. For example, cGMP-, calmodulin- and phospholipid-dependent phosphorylation have all been implicated in slow channel function. However, the role of protein phosphorylation in the regulation of channels during development is unknown. The proposed experiments seek to determine the role of protein phosphorylation during development of the embryonic chick heart. Changes in cAMP levels occur during development; whether cGMP levels also change is not known. Calmodulin levels, cGMP levels and various protein kinase activities will be measured at different stages of development. Regulation of sarcolemmal proteins various protein kinases will be examined to determine whether the pattern of phosphorylation changes during development, as has been shown for cAMP- dependent phosphorylation. The role of dephosphorylation will also be examined electrophysiologically and biochemically. To identify and further study the Ca channel protein(s) during development, a specific covalent affinity reagent, (3H)nifedipine isothiocyanate will be used to label the channel. Biochemical and electrophysiological experiments will be correlated to give a better understanding of channel properties and function during development. These studies are improtant, since the response of the heart to pathological conditions (e.g., myocardial ischemia) and to cardioactive drugs is largely determined by the types and properties of channels present.